Ever-young sex chromosomes in European tree frogs.

Non-recombining sex chromosomes are expected to undergo evolutionary decay, ending up genetically degenerated, as has happened in birds and mammals. Why are then sex chromosomes so often homomorphic in cold-blooded vertebrates? One possible explanation is a high rate of turnover events, replacing master sex-determining genes by new ones on other chromosomes. An alternative is that X-Y similarity is maintained by occasional recombination events, occurring in sex-reversed XY females. Based on mitochondrial and nuclear gene sequences, we estimated the divergence times between European tree frogs (Hyla arborea, H. intermedia, and H. molleri) to the upper Miocene, about 5.4-7.1 million years ago. Sibship analyses of microsatellite polymorphisms revealed that all three species have the same pair of sex chromosomes, with complete absence of X-Y recombination in males. Despite this, sequences of sex-linked loci show no divergence between the X and Y chromosomes. In the phylogeny, the X and Y alleles cluster according to species, not in groups of gametologs. We conclude that sex-chromosome homomorphy in these tree frogs does not result from a recent turnover but is maintained over evolutionary timescales by occasional X-Y recombination. Seemingly young sex chromosomes may thus carry old-established sex-determining genes, a result at odds with the view that sex chromosomes necessarily decay until they are replaced. This raises intriguing perspectives regarding the evolutionary dynamics of sexually antagonistic genes and the mechanisms that control X-Y recombination.

Inhibition of toxic epidermal necrolysis by blockade of CD95 with human intravenous immunoglobulin.

Toxic epidermal necrolysis (TEN, Lyell's syndrome) is a severe adverse drug reaction in which keratinocytes die and large sections of epidermis separate from the dermis. Keratinocytes normally express the death receptor Fas (CD95); those from TEN patients were found to express lytically active Fas ligand (FasL). Antibodies present in pooled human intravenous immunoglobulins (IVIG) blocked Fas-mediated keratinocyte death in vitro. In a pilot study, 10 consecutive individuals with clinically and histologically confirmed TEN were treated with IVIG; disease progression was rapidly reversed and the outcome was favorable in all cases. Thus, Fas-FasL interactions are directly involved in the epidermal necrolysis of TEN, and IVIG may be an effective treatment.

Dopamine reverses reward insensitivity in apathy following globus pallidus lesions.

Apathy is a complex, behavioural disorder associated with reduced spontaneous initiation of actions. Although present in mild forms in some healthy people, it is a pathological state in conditions such as Alzheimer's and Parkinson's disease where it can have profoundly devastating effects. Understanding the mechanisms underlying apathy is therefore of urgent concern but this has proven difficult because widespread brain changes in neurodegenerative diseases make interpretation difficult and there is no good animal model. Here we present a very rare case with profound apathy following bilateral, focal lesions of the basal ganglia, with globus pallidus regions that connect with orbitofrontal (OFC) and ventromedial prefrontal cortex (VMPFC) particularly affected. Using two measures of oculomotor decision-making we show that apathy in this individual was associated with reward insensitivity. However, reward sensitivity could be established partially with levodopa and more effectively with a dopamine receptor agonist. Concomitantly, there was an improvement in the patient's clinical state, with reduced apathy, greater motivation and increased social interactions. These findings provide a model system to study a key neuropsychiatric disorder. They demonstrate that reward insensitivity associated with basal ganglia dysfunction might be an important component of apathy that can be reversed by dopaminergic modulation.

A critical lineage-nonspecific role for pTalpha in mediating allelic and isotypic exclusion in TCRbeta-transgenic mice.

Although it is well established that early expression of TCRbeta transgenes in the thymus leads to efficient inhibition of both endogenous TCRbeta and TCRgamma rearrangement (also known as allelic and "isotypic" exclusion, respectively) the role of pTalpha in these processes remains controversial. Here, we have systematically re-evaluated this issue using three independent strains of TCRbeta-transgenic mice that differ widely in transgene expression levels, and a sensitive intracellular staining assay that detects endogenous TCRVbeta expression in individual immature thymocytes. In the absence of pTalpha, both allelic and isotypic exclusion were reversed in all three TCRbeta-transgenic strains, clearly demonstrating a general requirement for pre-TCR signaling in the inhibition of endogenous TCRbeta and TCRgamma rearrangement. Both allelic and isotypic exclusion were pTalpha dose dependent when transgenic TCRbeta levels were subphysiological. Moreover, pTalpha-dependent allelic and isotypic exclusion occurred in both alphabeta and gammadelta T cell lineages, indicating that pre-TCR signaling can potentially be functional in gammadelta precursors. Finally, levels of endogenous RAG1 and RAG2 were not down-regulated in TCRbeta-transgenic immature thymocytes undergoing allelic or isotypic exclusion. Collectively, our data reveal a critical but lineage-nonspecific role for pTalpha in mediating both allelic and isotypic exclusion in TCRbeta-transgenic mice.

Altered Local Beta and Gamma Synchronization in Prefrontal Cortex of Mice with Redox Dysregulation or Maternal Immune Activation

Background: A developmental dysregulation of glutathione (GSH) synthesis leading to oxidative stress, when combined with environmental risk factors (viral infections) generating reactive oxygen species, can play a critical role in inducing schizophrenia phenotypes. GSH deficit induces morphological, physiological and behavioral anomalies analogous to those reported in schizophrenic patients, including disrupted parvalbumine (PV) inhibitory interneuron's integrity and neuronal synchrony (β/γ-oscillations). Methods: We assessed PV immunoreactivity (PV-IR) and local synchronization in prefrontal cortex of two mouse models: (1) mice with a genetic deficit in GSH (GCLM-/-) and (2) mice with prenatal immune activation at embryonic day17 (PolyI:C). Results: Adults from both mice models display reduced PV-IR in prefrontal cortex. In anterior cingulate (ACC) of GCLM-/-, appearance and maturation of PVI are delayed and worsened with peribubertal stress but not in adult one. This effect is reversed by treatment with the GSH precursor N-acetyl-cysteine. The power of beta and gamma oscillations are decreased in ACC of GCLM-/- while they increased in prelimbic cortex of PolyI:C mice. Conclusions: Despite reduced PV-IR in both models, alteration of the synchronization was different, indicating that the structural/functional disruption of the cortical circuitry was partly different in both models. Novel therapeutic strategies are proposed, based on interference with oxidative stress and inflammatory processes.

Bortezomib reverses the proliferative and antiapoptotic effect of neuropeptides on prostate cancer cells.

Objectives:  Neuropeptides are important signal initiators in advanced prostate cancer, partially acting through activation of nuclear factor kappa B. Central to nuclear factor kappa B regulation is the ubiquitin-proteasome system, pharmacological inhibition of which has been proposed as an anticancer strategy. We investigated the putative role of the proteasome inhibitor bortezomib in neuropeptides signaling effects on prostate cancer cells. Methods:  Human prostate cancer cell lines, LNCaP and PC-3, were used to examine cell proliferation, levels of proapoptotic (caspase-3, Bad) and cell cycle regulatory proteins (p53, p27, p21), as well as total and phosphorylated Akt and p44/42 mitogen-activated protein kinase proteins. Furthermore, 20S proteasome activity, subcellular localization of nuclear factor kappa B and transcription of nuclear factor kappa B target genes, interleukin-8 and vascular endothelial growth factor, were assessed. Results:  Neuropeptides (endothelin-1, bombesin) increased cell proliferation, whereas bortezomib decreased proliferation and induced apoptosis, an effect maintained after cotreatment with neuropeptides. Bad, p53, p21 and p27 were downregulated by neuropeptides in PC-3, and these effects were reversed with the addition of bortezomib. Neuropeptides increased proteasomal activity and nuclear factor kappa B levels in PC-3, and these effects were prevented by bortezomib. Interleukin-8 and vascular endothelial growth factor transcripts were induced after neuropeptides treatment, but downregulated by bortezomib. These results coincided with the ability of bortezomib to reduce mitogen-activated protein kinase signaling in both cell lines. Conclusions:  These findings are consistent with bortezomib-mediated abrogation of neuropeptides-induced proliferative and antiapoptotic signaling. Thus, the effect of the drug on the neuropeptides axis needs to be further investigated, as neuropeptide action in prostate cancer might entail involvement of the proteasome.

The evolution of XY recombination: sexually antagonistic selection versus deleterious mutation load.

Recombination arrest between X and Y chromosomes, driven by sexually antagonistic genes, is expected to induce their progressive differentiation. However, in contrast to birds and mammals (which display the predicted pattern), most cold-blooded vertebrates have homomorphic sex chromosomes. Two main hypotheses have been proposed to account for this, namely high turnover rates of sex-determining systems and occasional XY recombination. Using individual-based simulations, we formalize the evolution of XY recombination (here mediated by sex reversal; the "fountain-of-youth" model) under the contrasting forces of sexually antagonistic selection and deleterious mutations. The shift between the domains of elimination and accumulation occurs at much lower selection coefficients for the Y than for the X. In the absence of dosage compensation, mildly deleterious mutations accumulating on the Y depress male fitness, thereby providing incentives for XY recombination. Under our settings, this occurs via "demasculinization" of the Y, allowing recombination in XY (sex-reversed) females. As we also show, this generates a conflict with the X, which coevolves to oppose sex reversal. The resulting rare events of XY sex reversal are enough to purge the Y from its load of deleterious mutations. Our results support the "fountain of youth" as a plausible mechanism to account for the maintenance of sex-chromosome homomorphy.

The early Miocene rotation of the Corso-Sardinian block. New paleomagnetic constraints for the end of the motion

The paleomagnetic investigations carried out in the 70's on Oligo-Miocene volcanics of Sardinia have demonstrated that the island was turned by 35-30 degrees clockwise from 33 Ma up to 3-1-20.5 Ma and rotated counterclockwise in a few million years [De Jong et al., 1969, 1973; Bobier et Coulon, 1970; Coulon et al., 1974; Manzoni, 1974, 1975; Bellon rr nl.. 1977: Edel et Lortscher, 1977; Edel, 1979, 1980]. Since then, the end of the rotation fixed at 19 Ma by Montigny er al. [1981] was the subject of discussions and several studies associating paleomagnetism and radiometric dating were undertaken [Assorgia er al., 1994: Vigliotti et Langenheim, 1995: Deino et al., 1997; Gattacceca rt Deino, 1999]. This is a contribution to this debate that is hampered by thr important secular variation recorded in the volcanics. The only way to get our of this problem is to sample series of successive flows as completely as possible, and to reduce the effect of secular variation by the calculation of means. Sampling was performed north of Bonorva in 5 pyroclastic flows that belong to the upper ignimbritic series SI2 according to Coulon rr nl. [1974] or LBLS according to Assorgia et al. [1997] (fig. I). Ar-40/Ar-39 dating of biotites from the debris flow (MDF) has yielded an age or 18.35 +/- 0.03 Ma [Dubois, 2000]. Five of the investigated sites are located beneath the debris flow ITV, TVB, TVD, SPM85, SPM86), one site was cured in the matrix of the debris flow (MDF) and one in 4 metric blocks included in the flow (DFC). Another site was sampled in the upper ash flow (PDM) that marks the end of the pyroclastic activity, just before the marine transgression. According to micropaleontological and radiometric dating this transgression has occurred between 18.35 and 17.6 Ma [Dubois, 2000]. After removal of a soft viscous component, the thermal demagnetization generally shows a univectorial behaviour of the remanent magnetization (fig. 2a). The maximum unblocking temperatures of 580-620 degrees (tab. I) and a rapid saturation below 100 mT (fig. 3) indicate that the carrier of the characteristic magnetization is magnetite. The exception comes: from the upper site PDM in which were found two characteristic components, one with a normal polarity and low unblocking temperatures up to 350 degreesC and one with a reversed polarity and maximum unblocking temperatures at 580-600 degreesC of magnetite. After calculation of a mean direction for each flow, the mean << Al >> direction 4 degrees /57 degrees (alpha (95) = 13 degrees) computed with the mean directions for the 5 flows may be considered as weakly affected by secular variation. But the results require a more careful examination. The declinations are N to NNW beneath the debris flow. NNW in the debris flow. and NNE (or SSW) above the debris flow, The elongated distribution of the directions obtained at sites TVB and TVD. scattered from the mean direction of TV to the mean direction of MDF is interpreted as due to partial overprinting during the debris How volcanic episode, The low temperature component PDMa is likely related to the alteration seen on thin sections and is also viewed as an overprint. As NNE/SSW directions occur as well below (mean direction << B >> : 5 degrees /58 degrees) as above the debris flow (PDMb : 200 degrees/-58 degrees). the NNW directions (<< C >> : 337 degrees /64 degrees) associated with the debris flow volcanism may be interpreted as resulting from a magnetic field excursion. According to the polarity scale of Cande and Kent [1992, 1995] and the radiometric age of MDF, the directions with normal polarity (TV, TVB, TVD, SPM85. SPM86a. MDF. DFC) may represent the period 5En. while the directions with reversed polarity PDMb and SPM86b were likely acquired during the period 5Dr. Using the mean << Al >> direction, the mean << B >>, or the PDM direction (tab. I). the deviation in declination with the direction of stable Europe 6.4 degrees /58.7 degrees (alpha (95) = 8 degrees) for a selection of 4 middle Tertiary poles by Besse et Courtillot [1991] or 7 degrees /56 degrees (alpha (95) = 3 degrees) for 19 poles listed by Edel [1980] can be considered as negligible. Using the results from the uppermost ignimbritic layer of Anglona also emplaced around 18.3 Ma [Odin rt al.. 1994]. the mean direction << E >> (3 degrees /51.5 degrees) leads to the same conclusion. On the contrary, when taking into account all dated results available for the period 5En (mean direction << D >> 353 degrees /56 degrees for 45 sites) (tab. II). the deviation 13 degrees is much more significant. As the rotation of Sardinia started around 21-20.5 Ma. the assumption of a constant velocity of rotation and the deviations of the Sardinia directions with respect to the stable Europe direction locate the end of the motion between 18.3 and 17.2 or 16.7 Ma (fig. 4). During the interval 18.35-17.5 Ma, the marine transgression took place. At the same period a NE-SW shortening interpreted as resulting from the collision of Sardinia with Apulia affected different parts of the island [Letouzey et al., 1982]. Consequently, the new paleomagnetic results and the tectono-sedimentary evolution are in favour of an end of the rotation at 17.5-18 Ma.

Delta9-tetrahydrocannabinol decreases somatic and motivational manifestations of nicotine withdrawal in mice

The possible interactions between Delta9-tetrahydrocannabinol (THC) and nicotine remain unclear in spite of the current association of cannabis and tobacco in humans. The aim of the present study was to explore the interactions between these two drugs of abuse by evaluating the consequences of THC administration on the somatic manifestations and the aversive motivational state associated to nicotine withdrawal in mice. Acute THC administration significantly decreased the incidence of several nicotine withdrawal signs precipitated by mecamylamine or naloxone, such as wet-dog-shakes, paw tremor and scratches. In both experimental conditions, the global withdrawal score was also significantly attenuated by acute THC administration. THC also reversed conditioned place aversion associated to naloxone precipitated nicotine withdrawal. We have then evaluated whether this effect of THC was due to possible adaptive changes induced by chronic nicotine on CB1 cannabinoid receptors. The stimulation of GTPS-binding proteins by the cannabinoid agonist WIN 55,212-2 and the density of CB1 cannabinoid receptor binding labelled with [3H] CP-55,940 were not modified by chronic nicotine treatment in the different brain structures investigated. Finally, we evaluated the consequences of THC administration on c-Fos expression in several brain structures after chronic nicotine administration and withdrawal. c-Fos was decreased in the caudate putamen and the dentate gyrus after mecamylamine precipitated nicotine withdrawal. However, acute THC administration did not modify c-Fos expression under these experimental conditions. Taken together, these results indicate that THC administration attenuated somatic signs of nicotine withdrawal and this effect was not associated to compensatory changes on CB1 cannabinoid receptors during chronic nicotine administration. In addition, THC also ameliorated the aversive motivational consequences of nicotine withdrawal.

BDNF impairment in the hippocampus is related to enhanced despair behavior in CB1 knockout mice

Stress can cause damage and atrophy of neurons in the hippocampus by deregulating the expression of neurotrophic factors that promote neuronal plasticity. The endocannabinoid system represents a physiological substrate involved in neuroprotection at both cellular and emotional levels. The lack of CB1 receptor alters neuronal plasticity and originates an anxiety-like phenotype in mice. In the present study, CB1 knockout mice exhibited an augmented response to stress revealed by the increased despair behavior and corticosterone levels showed in the tail suspension test and decreased brain derived neurotrophic factor (BDNF) levels in the hippocampus. Interestingly, local administration of BDNF in the hippocampus reversed the increased despair behavior of CB1 knockout mice, confirming the crucial role played by BDNF on the emotional impairment of these mutants. The neurotrophic deficiency seems to be specific for BDNF since no differences were found in the levels of NGF and NT-3, two additional neurotrophic factors. Moreover, BDNF impairment is not related to the activity of its specific receptor TrkB or the activity of the transcription factor CREB. These results suggest that the lack of CB1 receptor originates an enhanced response to stress and neuronal plasticity by decreasing BDNF levels in the hippocampus that lead to impairment in the responses to emotional disturbances.

A short motif in Drosophila SECIS Binding Protein 2 provides differential binding affinity to SECIS RNA hairpins

Selenoproteins contain the amino acid selenocysteine which is encoded by a UGA Sec codon. Recoding UGA Sec requires a complex mechanism, comprising the cis-acting SECIS RNA hairpin in the 3′UTR of selenoprotein mRNAs, and trans-acting factors. Among these, the SECIS Binding Protein 2 (SBP2) is central to the mechanism. SBP2 has been so far functionally characterized only in rats and humans. In this work, we report the characterization of the Drosophila melanogaster SBP2 (dSBP2). Despite its shorter length, it retained the same selenoprotein synthesis-promoting capabilities as the mammalian counterpart. However, a major difference resides in the SECIS recognition pattern: while human SBP2 (hSBP2) binds the distinct form 1 and 2 SECIS RNAs with similar affinities, dSBP2 exhibits high affinity toward form 2 only. In addition, we report the identification of a K (lysine)-rich domain in all SBP2s, essential for SECIS and 60S ribosomal subunit binding, differing from the well-characterized L7Ae RNA-binding domain. Swapping only five amino acids between dSBP2 and hSBP2 in the K-rich domain conferred reversed SECIS-binding properties to the proteins, thus unveiling an important sequence for form 1 binding.

THC prevents MDMA neurotoxicity in mice

The majority of MDMA (ecstasy) recreational users also consume cannabis. Despite the rewarding effects that both drugs have, they induce several opposite pharmacological responses. MDMA causes hyperthermia, oxidative stress and neuronal damage, especially at warm ambient temperature. However, THC, the main psychoactive compound of cannabis, produces hypothermic, anti-inflammatory and antioxidant effects. Therefore, THC may have a neuroprotective effect against MDMA-induced neurotoxicity. Mice receiving a neurotoxic regimen of MDMA (20 mg/kg ×4) were pretreated with THC (3 mg/kg ×4) at room (21°C) and at warm (26°C) temperature, and body temperature, striatal glial activation and DA terminal loss were assessed. To find out the mechanisms by which THC may prevent MDMA hyperthermia and neurotoxicity, the same procedure was carried out in animals pretreated with the CB1 receptor antagonist AM251 and the CB2 receptor antagonist AM630, as well as in CB1, CB2 and CB1/CB2 deficient mice. THC prevented MDMA-induced-hyperthermia and glial activation in animals housed at both room and warm temperature. Surprisingly, MDMA-induced DA terminal loss was only observed in animals housed at warm but not at room temperature, and this neurotoxic effect was reversed by THC administration. However, THC did not prevent MDMA-induced hyperthermia, glial activation, and DA terminal loss in animals treated with the CB1 receptor antagonist AM251, neither in CB1 and CB1/CB2 knockout mice. On the other hand, THC prevented MDMA-induced hyperthermia and DA terminal loss, but only partially suppressed glial activation in animals treated with the CB2 cannabinoid antagonist and in CB2 knockout animals. Our results indicate that THC protects against MDMA neurotoxicity, and suggest that these neuroprotective actions are primarily mediated by the reduction of hyperthermia through the activation of CB1 receptor, although CB2 receptors may also contribute to attenuate neuroinflammation in this process.

In vitro effect of malachite green on Candida albicans involves multiple pathways and transcriptional regulators UPC2 and STP2.

In this study, we show that a chemical dye, malachite green (MG), which is commonly used in the fish industry as an antifungal, antiparasitic, and antibacterial agent, could effectively kill Candida albicans and non-C. albicans species. We have demonstrated that Candida cells are susceptible to MG at a very low concentration (MIC that reduces growth by 50% [MIC(50)], 100 ng ml(-1)) and that the effect of MG is independent of known antifungal targets, such as ergosterol metabolism and major drug efflux pump proteins. Transcriptional profiling in response to MG treatment of C. albicans cells revealed that of a total of 207 responsive genes, 167 genes involved in oxidative stress, virulence, carbohydrate metabolism, heat shock, amino acid metabolism, etc., were upregulated, while 37 genes involved in iron acquisition, filamentous growth, mitochondrial respiration, etc., were downregulated. We confirmed experimentally that Candida cells exposed to MG resort to a fermentative mode of metabolism, perhaps due to defective respiration. In addition, we showed that MG triggers depletion of intracellular iron pools and enhances reactive oxygen species (ROS) levels. These effects could be reversed by the addition of iron or antioxidants, respectively. We provided evidence that the antifungal effect of MG is exerted through the transcription regulators UPC2 (regulating ergosterol biosynthesis and azole resistance) and STP2 (regulating amino acid permease genes). Taken together, our transcriptome, genetic, and biochemical results allowed us to decipher the multiple mechanisms by which MG exerts its anti-Candida effects, leading to a metabolic shift toward fermentation, increased generation of ROS, labile iron deprivation, and cell necrosis.

Poly-N-Acetyl Glucosamine Fibers Induce Angiogenesis in ADP Inhibitor-Treated Diabetic Mice

Background: It has been previously demonstrated that short-fiber poly-N-acetyl-glucosamine (sNAG) nanofibers specifically interact with platelets, are hemostatic, and stimulate diabetic wound healing by activating angiogenesis, cell proliferation, and reepithelialization. Platelets play a significant physiologic role in wound healing. The influence of altered platelet function by treatment with the ADP inhibitor Clopidogrel (CL) on wound healing and the ability of sNAG to repair wounds in diabetic mice treated with CL were studied.Methods: Dorsal 1 cm2 skin wounds were excised on genetically diabetic 8-week to 12-week-old, Lep/r-db/db male mice, and wound healing kinetics were determined. Microscopic analysis was performed for angiogenesis (PECAM-1) and cell proliferation (Ki67). Mice were either treated with CL (P2Y12 ADP receptor antagonist, CL) or saline solution (NT). CL wounds were also treated with either a single application of topical sNAG (CL-sNAG) or were left untreated (CL-NT).Results: CL treatment did not alter wound healing kinetics, while sNAG induced faster wound closure in CL-treated mice compared with controls. CL treatment of diabetic mice caused an augmentation of cell proliferation and reduced angiogenesis compared with nontreated wounds. However, sNAG reversed the effects of CL on angiogenesis and partially reversed the effect on cell proliferation in the wound beds. The sNAG-treated wounds in CL-treated mice showed higher levels of cell proliferation and not did inhibit angiogenesis.Conclusions: CL treatment of diabetic mice decreased angiogenesis and increased cell proliferation in wounds but did not influence macroscopic wound healing kinetics. sNAG treatment did not inhibit angiogenesis in CL-treated mice and induced faster wound closure; sNAG technology is a promising strategy to facilitate the healing of complex bleeding wounds in CL-treated diabetic patients.